TRANSCRIPTION OF HERPES SIMPLEX VIRUS GLYCOPROTEIN GENES

单纯疱疹病毒糖蛋白基因的转录

• 批准号: 3131494
• 负责人: LEWIS I PIZER
• 金额: $ 12.99万
• 依托单位: UNIVERSITY OF COLORADO DENVER
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-07-01 至 1989-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3131494
• 关键词: Alphaherpesvirinae; DNA directed RNA polymerase; HeLa cells; gel filtration chromatography; genetic manipulation; genetic recombination; genetic transcription; messenger RNA; monoclonal antibody; mutant; nucleic acid sequence; plasmids; radiotracer; regulatory gene; thymidine kinase; tissue /cell culture; virus DNA; virus genetics; virus infection mechanism

Herpes simplex virus proteins synthesized in infected cells change in both number and character as the infection proceeds. Underlying these changes in viral proteins are changes in the species and abundance of the viral mRNAs. We proposed that the time of appearance of early and late HSV mRNA is regulated predominantly by the ability to initiate transcription and that several factors control this process. These include primarily DNA sequences upstream from the promotor and viral regulatory proteins. Our goal is to identify the regulatory proteins and the DNA sequences with which they interact. To assay for upstream control elements in viral DNA we will construct recombinant plasmids containing isolated HSV genes, modify them by mutational insertions or deletions and assay their activity in vitro and in vivo. To study transcriptional regulation from these genes, we will use a three-pronged approach that measures RNA synthesis in nuclear extracts, in isolated nuclei, and in cells following transfection with plasmid DNA. The results from the in vivo experiments will be used to assess the validity of the in vitro measurements. Taken together, the data will establish whether transcription of HSV genes is regulated by viral proteins acting either independently or in a coordinate manner and whether specific gene sequences are involved. Long term goals are to study the biochemical interactions between the viral proteins and the enzymes involved in RNA synthesis. The availability of an in vitro transcription system makes the biochemical approach possible.

感染细胞中合成的单纯疱疹病毒蛋白 随着感染的进行，数量和特征都会发生变化。 病毒蛋白的这些变化背后是 病毒mRNA的种类和丰度。我们建议， HSV mRNA出现早、晚的时间受调控 主要是通过启动转录的能力和 有几个因素控制着这一过程。这些措施主要包括 启动子上游的DNA序列和病毒调控 蛋白质。 我们的目标是识别调控蛋白和DNA 它们相互作用的序列。对上游进行检测 我们将构建重组病毒DNA中的控制元件 含有分离的HSV基因的质粒，通过以下方式修改它们 突变插入或缺失并检测其活性 体外和体内。研究转录调控 这些基因，我们将使用三管齐下的方法来测量 在核提取液中、在分离的细胞核中和在 用质粒DNA转染后的细胞。 活体实验的结果将用于评估 体外测量的有效性。总而言之， 数据将确定HSV基因的转录是否 由病毒蛋白独立作用或在 协调方式和特定基因序列是否 牵涉其中。长期目标是研究生物化学 病毒蛋白与酶的相互作用 参与了RNA的合成。阿司匹林的体外可利用性 转录系统使生化方法成为可能。